BD® OMICS-One Immuno-Oncology Protein Panel

Reactivity: Human (Tested in Development)
Application: Single Cell 3' Sequencing, Single Cell 3' Sequencing (Qualified)
Regulatory Status: RUO
Description: Description The BD® OMICS-One Immuno-Oncology Protein Panel consists of six single tubes: two tubes of T-Cell Protein Panel (1 test/tube) containing 30 different specificities against major T-cell markers, two tubes of B-Cell Protein Panel (1 test/tube) containing 30 different specificities against major B-cell markers, and two tubes of Tumor Protein Panel (1 test/tube) containing 30 different specificities against major Tumor markers. Designed and optimized to work on the BD Rhapsody™ System, the Immuno-Oncology Protein Panel is tested to work seamlessly alongside the BD Rhapsody™ Whole Transcriptome Analysis (WTA) Assay, Targeted mRNA Assay, BD® Single-Cell Multiplexing Kit (SMK), BD® Intracellular CITE-seq (IC-AbSeq) Assay, and BD Rhapsody™ TCR/BCR Next Multiomic Assay for human. The individual antibodies were each conjugated to an oligonucleotide that contains a specific antibody barcode sequence flanked by a polyA tail on the 3' end and a common PCR handle (PCR primer binding site) on the 5' end. All AbSeq barcode sequences were generated in silico with minimal sequence similarity to the human genomes, have low predicted secondary structure, and have high Hamming distance within the BD antibody-oligo portfolio, to allow for sequencing error correction and unique mapping. The polyA tail of the oligonucleotide allows the barcode sequence to be captured by the BD Rhapsody™ Enhanced Cell Capture Beads. The 5' PCR handle allows for efficient library generation for various sequencing platforms. Each individual antibody exists at an optimal concentration within the 59-plex panel to enable superior target and population resolution. The Immuno-Oncology Protein Panel is designed with SMART technology. SMART technology helps lower sequencing cost while increasing data resolution by attenuating antibodies that target high-expressing primary markers and allowing reallocation of sequencing reads to markers expressed at lower levels. With SMART technology, markers low in expression can be quantified without having to do deeper sequencing and incurring high sequencing cost. There are six specificities attenuated in the Immuno-Oncology Protein Panel: CD4, CD43, CD44, CD45, HLA-DR, and HLA-ABC.
Preparation And Storage: Preparation And Storage Store at 2-8°C and protected from prolonged exposure to light. Do not freeze.
Recommended Assay Procedures: Recommended Assay Procedures Note : The protocol supports the Ab-Oligos labeling of 1 − 2 million cells per tube. 1. Remove one tube of BD® OMICS-One T-Cell Protein Panel, one tube of BD® OMICS-One B-Cell Protein Panel, and one tube of BD® OMICS-One Tumor Protein Panel from foil bags and bring up to room temperature for 5 minutes. 2. Make sure all pellets are located at the bottom of the tubes. If not, briefly centrifuge to collect the contents at the tube bottom. 3. For each tube, add 35 µL of nuclease-free water to the bottom of the tube and allow antibodies to reconstitute for 5 minutes at room temperature. 4. Transfer the reconstituted antibodies on ice until the cells are ready for staining. Note: Reconstitute antibody immediately before cell staining. Prolonged incubation of reconstituted antibody may increase the non-specific background. 5. For BD® AbSeq Ab-Oligo drop-in of 25 plex or lower, prepare the BD® AbSeq labeling MasterMix in 1.5-mL LoBind tube on ice. The following calculation is for the procedure with Optional Fc Block. If Fc Block is not performed, add 25 µL BD Pharmingen™ Stain Buffer (FBS) to the Abseq labeling MasterMix total volume for each sample. Note: For drop-in with more than 25 plex, reach out to technical support for the calculation. For sequential labeling with Sample Tags or no Sample Tags, prepare BD® AbSeq labeling MasterMix for drop-ins as follows: ____________________________________________________________________________________________ Component                           1 sample (µL)       1 sample +          2 samples + 30% overage (µL)    30% overage (µL) Per BD® AbSeq Ab-Oligo              2.0                 2.6                 5.2 Total of BD® AbSeq Ab-Oligo         2.0 × N*            2.6 × N             5.2 × N FBS † (catalog number 554656)        70 – (2.0 x N)      91 – (2.6 x N)      182 – (5.2 x N) Total                               70                  91                  182 For co-labeling with Sample Tags, prepare BD® AbSeq labeling MasterMix for drop-ins as follows: ____________________________________________________________________________________________ Component                           1 sample (µL)       1 sample +          2 samples + 30% overage (µL)    30% overage (µL) Per BD® AbSeq Ab-Oligo              2.0                 2.6                 5.2 Total of BD® AbSeq Ab-Oligo         2.0 × N*            2.6 × N             5.2 × N FBS † (catalog number 554656)        50 – (2.0 × N)      65 – (2.6 × N)      130 – (5.2 × N) Total                               50                  65                  130 *  N = number of drop-in antibodies. N = 0 if there are no drop-in antibodies. †  FBS = BD Pharmingen™ Stain Buffer. 6. Pipet-mix the BD® AbSeq labeling MasterMix for drop-ins. Briefly centrifuge to collect the contents at the bottom, and place back on ice. 7. For sequential labeling with Sample Tags or no Sample Tags, for each sample, combine the three tubes containing 35 µL reconstituted T-Cell Protein Panel solution, 35 µL reconstituted B-Cell Protein Panel solution, and 35 µL reconstituted Tumor Protein Panel solution. Then add 70 µL BD® AbSeq labeling MasterMix of drop-ins to the tube containing 105 µL reconstituted T-Cell, B-Cell, and Tumor Protein Panel solution to make a total volume of 175 µL. For co-labeling with Sample Tags, for each sample, combine the three tubes containing 35 µL reconstituted T-Cell Protein Panel solution, 35 µL reconstituted B-Cell Protein Panel solution, and 35 µL reconstituted Tumor Protein Panel solution. Then add 50 µL BD® AbSeq labeling MasterMix of drop-ins and 20 µL Sample Tag to the tube containing total 105µL reconstituted T-Cell, B-Cell, and Tumor Protein Panel solution to make a total volume of 175 µL. 8. Pipet-mix the mixture, briefly centrifuge to collect the contents at the tube bottom, and place back on ice. 9. Centrifuge cells at 400 × g for 5 minutes. If Fc Block is used, proceed to step 10. If Fc Block is not used. skip to step 11. 10. (Optional) For samples containing myeloid and B lymphocytes, BD Biosciences recommends blocking nonspecific Fc Receptor–mediated false-positive signals with Human BD Fc Block (catalog number 564220). a. To perform blocking, pipet the Fc Block MasterMix into a new 1.5-mL LoBind tube on ice: _________________________________________________________________________________ Component                           1 sample (µL)*    1 sample + 20% overage (µL) FBS† (catalog number 554656)        20.0              24.0 Fc Block‡ (catalog number 564220)   5.0               6.0 Total                               25.0              30.0 *  Sufficient for up to 1 million cells. To block more cells, adjust the volume. †  FBS = BD Pharmingen™ Stain Buffer. ‡ Fc Block =  BD Pharmingen™ Human BD Fc Block. b. Pipet-mix the Fc Block MasterMix and briefly centrifuge. Place on ice. c. Remove the supernatant from the cells without disturbing the pellet. d. Resuspend the cells in 25 µL of Fc Block MasterMix. e. Incubate the cells at room temperature (15°C to 25°C) for 10 minutes. f. Add 175 µL of BD® AbSeq labeling MasterMix from Step 8 into the cell suspension. Pipet-mix and proceed to Step 12. 11. Remove the supernatant from the cells without disturbing the pellet. Add 25 µL Stain Buffer (FBS) to the 175 µL of BD® AbSeq labeling MasterMix from Step 8 to make a total volume of 200 µL. Resuspend the cell pellet in 200 µL total volume. Pipet-mix. 12. Transfer the cells with BD® AbSeq labeling MasterMix into a new 5-mL polystyrene Falcon tube. 13. Stain the cells on ice for 30 minutes. 14. Add 3–4 mL Stain Buffer (FBS) to labelled cells and pipet-mix. 15. Centrifuge at 400 x g for 5 minutes. 16. Uncap the tube and invert to decant supernatant into biohazardous waste. Keep the tube inverted and gently blot on a lint-free wiper to remove residual supernatant from tube rim. 17. Repeat steps 14–16 twice more for a total of three washes. 18. Resuspend the final washed cell pellet in 620 µL cold Sample Buffer from the BD Rhapsody™ Enhanced Cartridge Reagent V3 (catalog number 667052) and proceed to single cell capture with on-cartridge washing described in substeps a–c. Refer to the BD Rhapsody™ HT Single-Cell Analysis System Single-Cell Capture and cDNA Synthesis Protocol (Doc ID 23-24252) or BD Rhapsody™ HT Xpress System Single-Cell Capture and cDNA Synthesis Protocol (Doc ID 23-24253) for additional details. Note: Perform on-cartridge washing after cell settling (8 minute incubation) as follows: a. At the protocol section of "Loading cells in BD Rhapsody™ 8-Lane Cartridge", after cell load, incubate the cartridge in the dark at room temperature for 8 minutes. b. Place the cartridge on the BD Rhapsody™ HT Xpress and perform the On-Cartridge Wash steps listed as follows: _____________________________________________________________ Material to load        Volume (µL) 1 lane      Pipette Mode Air                     380                     Prime/Wash Cold Sample Buffer      380                     Prime/Wash Air                     380                     Prime/Wash Cold Sample Buffer      380                     Prime/Wash c. (Optional) Perform the scanner step: Cell Load Scan, if using BD Rhapsody™ HT Single-Cell Analysis System Single-Cell Capture and cDNA Synthesis Protocol (Doc ID 23-24252). No need for 8-minute delay before scanning. Warning: All biological specimens and materials are considered biohazardous. Handle as if capable of transmitting infection and dispose using proper precautions in accordance with federal, state, and local regulations. Never pipette by mouth. Wear suitable protective clothing, eyewear, and gloves. List of all 30 Human AbSeq specificities included in the BD® OMICS-One T-Cell panel: _____________________________________________________________________________ Specificity     Clone      Oligo ID     BD® AbSeq Barcode Sequence CD103           Ber-ACT8   AHS0001      AAATAGTATCGAGCGTAGTTAAGTTGCGTAGCCGTT CD137           4B4-1      AHS0003      TGACAAGCAACGAGCGATACGAAAGGCGAAATTAGT CD45RA          HI100      AHS0009      AAGCGATTGCGAAGGGTTAGTCAGTACGTTATGTTG CD69            FN50       AHS0010      CAATAACGGGTCATAGTAAGTCGCGAGTAAGAGGGC CD278           DX29       AHS0012      ATAGTCCGCCGTAATCGTTGTGTCGCTGAAAGGGTT CD134 (OX40)    ACT35      AHS0013      GGTGTTGGTAAGACGGACGGAGTAGATATTCGAGGT CD279 (PD-1) † EH12.1     AHS0014      ATGGTAGTATCACGACGTAGTAGGGTAATTGGCAGT CD366 (TIM-3)   7D3        AHS0016      TAGGTAGTAGTCCCGTATATCCGATCCGTGTTGTTT CD223 (Lag3)    T47-530    AHS0018      CGGCATGAATTAGGCGAGACTTAGTATACGAGCTGG CD95 (Fas)      DX2        AHS0023      GGCCCGTTAGAGTTGGTATCCGTATGAAGGTTAGCT CD25            2A3        AHS0026      AGTTGTATGGGTTAGCCGAGAGTAGTGCGTATGATT CD127           HIL-7R-M21 AHS0028      AGTTATTAGGCTCGTAGGTATGTTTAGGTTATCGCG CD183           1C6/CXCR3  AHS0031      AAAGTGTTGGCGTTATGTGTTCGTTAGCGGTGTGGG CD4             SK3        AHS0032      TCGGTGTTATGAGTAGGTCGTCGTGCGGTTTGATGT CD196 (CCR6)    11A9       AHS0034      ACGTGTTATGGTGTTGTTCGAATTGTGGTAGTCAGT CD45RO          UCHL1      AHS0036      TGAGAGGTTATTGGGCGTATGACTTCGGTGATTGTG CD194 (CCR4)    1G1        AHS0038      AATATTAGTGGGTCCTCGCGTTGGCCGGTTGTTAGT CD62L           DREG-56    AHS0049      ATGGTAAATATGGGCGAATGCGGGTTGTGCTAAAGT CD272           J168-540   AHS0052      GTAGGTTGATAGTCGGCGATAGTGCGGTTGAAAGCT CD154           TRAP1      AHS0077      TAAGAGGTAAGTGCATTCGGGTATAGGCGTGATTTG CD357 (GITR)    V27-580    AHS0104      TCTGTGTGTCGGGTTGAATCGTAGTGAGTTAGCGTG CD28            L293       AHS0138      TTGTTGAGGATACGATGAAGCGGTTTAAGGGTGTGG TCRgd           11F2       AHS0142      CTCGTGGGTTAGGCTTGATCGTAGTTATGTATGGTT CD44 † L178       AHS0167      GTGATTGATTAGGACAGTTCGTTGCTTAGTAGTGGG TCR Va24-Ja18   6B11       AHS0175      TTCTGGTTCGGTTGAGCTACTAATTTCGTTGGATGG CD161 (KLRB1)   HP-3G10    AHS0205      TTTAGGACGATTAGTTGTGCGGCATAGGAGGTGTTC CD8             SK1        AHS0228      AGGACATAGAGTAGGACGAGGTAGGCTTAAATTGCT CD3             UCHT1      AHS0231      AGCTAGGTGTTATCGGCAAGTTGTACGGTGAAGTCG CD197 (CCR7)    2-L1-A     AHS0273      AATGTGTGATCGGCAAAGGGTTCTCGGGTTAATATG TIGIT           tgMab-2    AHS0411      AGAGGGTTTAGTCAAGGTCGTGCGTATAGTTCAGGT List of all 30 Human AbSeq specificities included in the BD® OMICS-One B-Cell Panel: _____________________________________________________________________________ Specificity     Clone       Oligo ID    BD® AbSeq Barcode Sequence CD20            2H7         AHS0008     TTGCTTGTTGCGCGTTAGAGAGTATGTCGGGAGATG CD275           2D3/B7-H2   AHS0011     GTTTATATGTACGACGCCCGGTTGACGAGTGGAAGT CD38            HIT2        AHS0022     GTCAACGATGGGTAGCGGTAGAAATAACGGAACTGG CD95            DX2         AHS0023     GGCCCGTTAGAGTTGGTATCCGTATGAAGGTTAGCT CD27            M-T271      AHS0025     TGTCCGGTTTAGCGAATTGGGTTGAGTCACGTAGGT CD19            SJ25C1      AHS0030     TAGTAATGTGTTCGTAGCCGGTAATAATCTTCGTGG HLA-DR          G46-6       AHS0035     TGTTGGTTATTCGTTAGTGCATCCGTTTGGGCGTGG CD185 (CXCR5)   RF8B2       AHS0039     AGGAAGGTCGATTGTATAACGCGGCATTGTAACGGC CD24 ‡ ML5         AHS0042     ACTTTGGGTTGAGCGCATGATTATTCGTGACACTTT CD80            L307.4      AHS0046     GAGGGTAACGGGTGTCCAAATATCGGCTGTGTAAGT CD5             UCHT2       AHS0047     ACGAAGCGAGCGAAGAACCTATGCGATTGAGTAAGT CD10            HI10a       AHS0051     CCTGTTTGATGCGTACGGAGATTTAGCGGATTTATG IgD             IA6-2       AHS0058     TGAGGGATGTATAGCGAGAATTGCGACCGTAGACTT IgG             G18-145     AHS0059     AGGTAGGTTATCGTAGGGTAGACTTAGCGGGCATTG CD184 (CXCR4)   12G5        AHS0060     CAGTGTTTAGAGCGGGTTGCATATGTCGTTTAGAGG CD34 ‡ 581         AHS0061     TGGGTGTATTACGGTTAGTTTATGCGCGAAGGTGTT CD21            B-ly4       AHS0074     GTATTCGCGTATTGTCAGTCGGTAGGGTTATGGTCT CD9             M-L13       AHS0082     GGGTTGTAAGTCGTCGGAAGTGTGAAGCGTATAGTG CD126           M5          AHS0096     AATGGTGAATCGCCCTAGCAAGTGGTATCGGAATCG CD30            BERH8       AHS0114     CCAGTGTAGATTGAGCCGTCGATTTAGTTAGCAGTG CD40            5C3         AHS0117     GGTGTAATTGGGCTAGAACGTATATGCGGTAAGGCG CD138           MI15        AHS0121     TAAGCTGCCGGTATTGGAAACGTATCGATCTATTGG CD79B           CB3-1       AHS0153     CATCATGAGTAGTTGCTTCGGCGAGTAGGTTTAATT CD22            HIB22       AHS0195     TGGTTCGTGACTGTATAGGCTTAGCTTAGGCAATTT IgM             G20-127     AHS0198     TTTGGAGGGTAGCTAGTTGCAGTTCGTGGTCGTTTC CD43            1G10        AHS0200     ATGGCGGATGGATTTGTCGGTGATATTGCTCTCGTT CD268 (BAFF-R)  11C1        AHS0206     TGTGAATGAGTTAAGCGTCGCGGATATGTAGAGCCT CD23            EBVCS-5     AHS0210     TTTGATGTGGGCGGGTTGTATTACGGTTTCGAGTCT CD73            AD2         AHS0216     AAAGTAGGGTCGATCAAGGGAGTTAACGGTAGCGCT CD1d            CD1d42      AHS0219     GTTAGGATTATTGACGTACCGAGTTAGGAGTGATTG List of all 30 Human AbSeq specificities included in the BD® OMICS-One Tumor panel: _____________________________________________________________________________ Specificity     Clone      Oligo ID     BD® AbSeq Barcode Sequence CD274 (PD-L1) MIH1       AHS0004      ATCGTAAGGCTCGTGGTTCGTAAGTAAGTTCGTATC CD279 (PD-1) † EH12.1     AHS0014      ATGGTAGTATCACGACGTAGTAGGGTAATTGGCAGT CD45            HI30       AHS0040      GTGCGAAATGGCGGAATGTTATCTGCGAATGTAGTC CD324 (E-cad)   67A4       AHS0041      GATATGAATGGGTTGCGGTGTAAAGTCGTAATGGTT CD24 ‡ ML5        AHS0042      ACTTTGGGTTGAGCGCATGATTATTCGTGACACTTT CD90            5E10       AHS0045      GACTATATGTACGGTGTTAATTCGGGATCCTGCGCT CD34 ‡ 581        AHS0061      TGGGTGTATTACGGTTAGTTTATGCGCGAAGGTGTT CD117           YB5.B8     AHS0064      GGATTAGTTGTCGTTATAGGGAGTGCGTTCTTAGCG HLA-A,B,C       G46-2.6    AHS0066      GATATGCATGGCGAGTAGGTAGAACGAAGCTTAGGT CD54            HA58       AHS0076      AAGAGAATATATGCGTGCGTTGTTAAGGGAATGCGT CD29            MAR4       AHS0080      TGGTAAGGTGGTTGCGAGTAAGTAGCGGTGAGTTGT CD47            B6H12      AHS0087      TGTTAGGTTCGACGTATTATGTGTAGATCCGCAAGG CD326 (EpCam)   EBA-1      AHS0089      TTGAGCGTAAAGTTGCGTCCGGTAATTGAGTTGCGT CD66            B1.1/CD66  AHS0094      GTCTGCGCAAGGTAAGCTAAGTAACGAAAGGGATCT CD133           W6B3C1     AHS0103      TTTGGTATTGGCACGGTTTGTAGCGAGTTGACGGTC CD26            M-A261     AHS0109      TGTAGGTTGCGCGGTTATTAGGGTATTATCGATCTG CD155           TX24       AHS0111      GCGGTGGATCGATGGGTATAGTTGGTAATTTGCGTC CD146           P1H12      AHS0127      AGGTTATTTAGGTGACGGTTGTATTGACGAGAGAGG C-MET           3D6        AHS0132      AGCGTGAGTTGTCGGTAGTTAATTATCGGAGAGTTT ITGRN BTA 7     FIB504     AHS0158      TTTCAGTTTGGTCGCAGTTAAGGTATCGTATGGGTC CD44 † L178       AHS0167      GTGATTGATTAGGACAGTTCGTTGCTTAGTAGTGGG PECAM1 (CD31)   WM59       AHS0170      CTAAGGGACGTAATTGAGTTTCGGTGATCGCAGTTT EphB2           2H9        AHS0176      TATTGCGGGTAGGATTTGTCTCGAAGCGTAGGTAGC Vista           MIH65.RMAB AHS0187      ATCAGGGAATCTCGGTAAGTTAAACGTGTATAGTGC PDPLN           LPMAB-17   AHS0192      TTTATGAGTATTACGTCTGTTGCGATTGTTGGCGGT NOTCH1          MHN1-519   AHS0214      CGTAGTAGGAGCGTGTTTCATCGGCATTATCGTTTG CD325 (n-Cad)   8C11       AHS0223      TAGGATGAGTTTCGTAAGTAAGGTAGTCGTATGGCT CD58            1C3        AHS0237      TTGGTGAGTATTGGTGCGTAGTATGCGGGATGTTTG EGFR            EGFR.1     AHS0241      ATATGATTGATGCGGGTTAGCCTACAGATTCGAGTT CD227 (MUC1)    HMFG2      AHS0247      AGTGCATGGTTAGTAGGTGTGAGTCGTTAGATATTC * T-Cell and B-Cell Protein Panels contain the same CD95 antibody. † T-Cell and Tumor Protein Panels contain the same CD279 (PD-1) and CD44 antibody. ‡ B-Cell and Tumor Protein Panels contain the same CD24 and CD34 antibody.
Product Notices: Product Notices Please refer to www.bdbiosciences.com/us/s/resources for technical protocols. Source of all serum proteins is from USDA inspected abattoirs located in the United States. The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots. Please refer to http://regdocs.bd.com to access safety data sheets (SDS). For U.S. patents that may apply, see bd.com/patents. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing. Follow state and local guidelines when disposing of hazardous waste. Go to https://abseq-ref-gen.genomics.bd.com/to access AbSeq reference files in FASTA format for bioinformatics analyses. This reagent is provided lyophilized in a pre-titrated format.
Description : Quantity/Size : Part Number : EntrezGene ID
N/A : null : N/A : N/A